Diseases such as age related macular degeneration (AMD), other retinal dystrophies, and glaucoma cause a large proportion of blindness worldwide. A critical barrier in treating these diseases is the inability to efficaciously, economically, and safely deliver drugs and gene therapy DNA constructs to the retina and the retinal pigment epithelium (RPE). Topical delivery by eye drops has been ineffective because of permeability and absorption barriers of the anterior segment of the eye. Thus, current delivery approaches for these diseases entail costly injections of the eye using viral vectors that in some clinical trials have demonstrated dangerous side effects. The broad, long-term objective of this project is to develop a safe, economical, and effective gene delivery approach using novel, nonviral sub-50 nanometer (s50) capsule technology that is cell-specific and traffics to the nucleus without endosomal entrapment. Our preliminary data indicate that these nanocapsules administered as topical eye drops onto the tear film of living rats result in delivery and expression of gene vectors in cells of the retina and RPE. Topical application resulting in DNA delivery to posterior ocular targets is surprising and potentially transformative. If brought to the clinic, it should provide safe and economical treatment for many blinding diseases. In this Phase 1 study, we propose to optimize the topical s50-capsule dosing regimen in terms of safety and efficacy using easily-assessed fluorescent plasmid DNA expression vectors. We then will compare expression levels and inflammatory markers of the optimized topical s50-capsule dosing regimen, with an ocular injection protocol that we have previously optimized. We will use the LCA2 mouse model of retinal degeneration. LCA2 mouse models are useful because they have severe vision deficits due to simple lack of a single gene product (RPE65), yet they have delayed morphological degeneration. Thus, successful delivery of replacement gene vector results in creation of visual function and prevention of degeneration. If the results of this Phase I study are encouraging, we will propose Phase II studies using the novel s50 capsule technology to deliver therapeutic RPE65 expression vectors to mice deficient in RPE65, and assess target gene expression and function, visual function, and morphology. PUBLIC HEALTH RELEVANCE: The relevance of this research to public health is potentially very high. A large proportion of blindness is caused by retina and retinal pigment epithelium (RPE) pathologies. Several therapeutic DNA constructs and drugs are effective treatments for these diseases in animal and cell culture models, but are of limited use in the clinic because they cannot be delivered to target tissue and cells efficiently and safely. The aim of this project is to develop a safe and effective way to deliver these treatments.